The present invention relates to the production of monoclonal antibodies; and, in particular, to hybrid cell lines capable of continuously producing monoclonal antibodies directed against the outer membrane antigens of the bacterium Haemophilus influenzae.
In recent years, the capability to produce monoclonal antibodies specific for the immunogenic determinants of bacterial cells and toxins has provided a new vista of diagnostic and immunotherapeutic agents.
Haemophilus influenzae type b (Hib) is a bacterial pathogen attributed as the leading cause of endemic bacterial meningitis in infants and young children. This organism is also responsible for a number of other serious diseases in children, including epiglottitis and pneumonia.
The gravity of Hib meningitis is reflected by a mortality rate of 5-10%, despite treatment with antibiotics. Furthermore, a sizable percentage of survivors of Hib meningitis exhibit serious neurological sequelae. This situation, coupled with the recent emergence of ampicillin resistant strains of Hib, indicates the necessity for development of an effective vaccinogen to prevent Hib disease.
The currently available Hib vaccine featuring purified phosphoribosylribitol phosphate (PRRP) capsular antigen is not effective in inducing the formation of protective antibodies in children vaccinated at less than 14 months of age. Unfortunately, this group of children represents those persons at highest risk for systemic Hib disease.
Heretofore, it was generally assumed that serum antibody directed against the PRRP capsular element of Hib was responsible for resistance to Hib disease in humans. Recently, however, the protective value of anti-capsular antibody has been questioned, prompting the suggestion that antibody directed against somatic non-capsular Hib antigens may be equally or more important in providing resistance to Hib disease. In experiments designed to test prospective vaccinogens, infant rats which respond poorly to PRRP antigen mount a significant antibody response to general somatic non-capsular Hib antigens. However, the identity of the Hib cell surface immunogens to which these protective antibodies were directed had not been established prior to Applicant's work.
Two classes of Hib cell surface antigens against which protective antibodies might be directed include the lipopolysaccarides and proteins present in the outer membrane of the Hib pathogen. The inherent toxicity of most bacterial lipopolysaccharide molecules weighs against the implementation of this moiety as a potential Hib vaccinogen. In contrast, proteinaceous vaccines such as the diphtheria/pertussis/tetanus vaccine are relatively non-toxic and immunogenic in infants. Accordingly, the outer membrane proteins of Hib provide potential as a preferred agent for vaccinogens.
Conclusive proof that antibodies directed against Hib outer membrane antigens protect against systemic Hib disease requires the use of antibodies specific for these antigens. The recent development of lymphocyte hybridoma technology has made possible the production of monoclonal antibodies specific for any given antigen. While monoclonal antibodies have been extensively employed and manipulated in immunology, virology and parasitology research, research and application potential of monoclonal antibodies has only recently been tapped for microbial pathogenesis investigations. For example, monoclonal antibodies have been available for sometime now which are specific for an assortment of antigens, including viral antigens, such as rabies, hepatitis and influenza virus; red blood cells; fluorescent dyes; and cell associated antigens. More recently, monoclonal antibodies have been directed against bacterial components including different streptococcal antigens described by Briles et al, J. Exp. Med. 153:694-705 (1981) and Polin et al, J. Clin. Microbiol. 11:332-336 (1980); and cell surface antigens of Neisseria gonorrhoeae reported by Nachamkin et al, Infect. Immun. 32:641-648 (1981) and Johnston et al, Abstr. Ann. Meet. Am. Soc. Microbiol. B86, p. 29 (1981). Heretofore, as far as Applicants are aware, there have been no reports of the production of continuous cell lines of somatic cell hybrids which elaborate monoclonal antibodies to Hib outer membrane antigens, and in particular, to proteinaceous cell surface-exposed antigens.
It therefore is highly desirable to provide a means for producing antibody to outer membrane antigens of Hib. Such antibodies would be important in the diagnosis of Hib disease in humans, in the purification of specific immunogens for subsequent use as vaccines, and in use as highly specific immunotherapeutic agents to confer passive immunity on a host in the event of Hib infection.